gp120 HIV envelope glycoprotein increases the production of nitric oxide in human monocyte-derived macrophages

The effect of recombinant gp120 HIV envelope glycoprotein on the generation of free radicals by monocyte-derived macrophages (MDM) was measured by EPR spin trapping with 5,5-dimethyl-1-pyrroline-N-oxide (DMPO). After 1 day in culture, MDM produced a spin trap adduct of DMPO with hyperfine splitting constants superimposable on those of DMPO-OH. The addition of gp120 to MDM increased the production of DMPO-OH and after 1 h, the amount of DMPO-OH produced by 40 micrograms/ml gp120 was about 300% that of untreated MDM. The use of selective inhibitors suggested the participation of the nitric oxide/L-arginine oxidative pathway, but did not provide evidence for trapping of hydroxyl radical or other oxygen free radicals. The specificity of gp120 was proven by two different anti-gp120 antibodies that either inhibited (polyclonal) or increased (monoclonal) the production of free radicals. Dexamethasone inhibited the effect of gp120, suggesting the possible involvement of an inducible nitric oxide (NO) synthase. Moreover, treatment of MDM with gp120 for 15 h increased in a dose-dependent manner the production of NO2-, a stable end product of NO. Soluble CD4 did not modify the intensity of the DMPO-OH adduct, whereas yeast mannan and Ca(2+)-chelators abolished the increase in the DMPO-OH signal induced by gp120. These data suggest the possible involvement of mannose-specific endocytotic lectin of MDM. The reaction of DMPO with sodium nitroprusside, an organic nitrate that releases NO, also produced DMPO-OH. Our findings indicate that gp120 increases free radical production from MDM as detected by spin-trapping methods, and that the spin trap adduct results from a reaction involving NO or closely related oxidized derivatives.